polarity check

Submitted by AndreasSchmidt on Thu, 2016-07-21 04:56

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Frealix

Hi Alexis,

this is probably a stupid question, but how does Frealix compare different filaments in order to check if they are in the same orientation? How does Frealix know if fibrils are polar or not, so how does the polarity check work?

Thank you very much and all the best,

Andreas

Alexis Thu, 2016-07-21 09:39

Hi Andreas,

Most filaments are polar. Only those with a side dyad (a symmertry axis perpendicular to the helical axis), like DNA, are not.

When Frealix checks the polarity of the filament, it compares the best alignment it can get of the filament in the current direction versus the best alignment it can get when the order of waypoints and the direction of the filament is reversed. Of course, this will only work if the 3D map used is polar.

I hope this helps.

Alexis

AndreasSchmidt Tue, 2016-07-26 03:04

Hi Alexis,

thank you very much. Now I understand a bit better, how Frealix decides to flip the orientation of a fibril or to keep it.
So in this case, Frealix will use the single fibril reconstructions rather than the fibril images on the micrograph?

Best,

Andreas

Alexis Tue, 2016-07-26 08:38

Hi Andreas,

I'm not sure whether I understood your question correctly. Maybe this answers it:

The polarity check is done by using the 3D reconstruction from the previous cycle of refinement (i.e. the consensus reconstruction, with all filaments, called frealix_round_???/3drec.mrc) and correlating it against the fibril image on the micrograph. The correlation is done in both possible polarities, and only the best polarity is kept.

Hope this clarifies it.

Alexis

AndreasSchmidt Tue, 2016-07-26 09:10

Hi Alexis,

now I got it. This makes much more sense to me now.
I thought the fibril on the micrograph will be cut out and compared with itself after rotating it 180° (or the single fibril reconstructions) to see if they are polar, which would not make much sense because you only get the information if they are polar or not without knowing the orientation compared to the other ones.

Best,
Andreas